Determination of the Promoter Activity of HIV-1 Using the Chloramphenicol Acetyltransferase Reporter Gene Assay

RNA transcripts are produced from the 5′ long terminal repeat (LTR) of human immunodeficiency virus type-1 (HIV-1) under the complex control of viral regulatory sequences ( 1 – 7 ). The LTR from a prototypical, laboratory-adapted HIV-1 (HXB2) can be divided into modulatory, core promoter, and transactivating regions (Fig. 1 ). The modulatory regions include elements with limited homology to AP-1 enhancer sequences at positions −347 to −329 relative to the cap site ( 8 ), two NF-AT sites at positions −292 to −255 ( 9 ), a USF site at positions −159 to −173 ( 10 – 12 ), a TCF-lα site at positions −139 to −124 ( 13 ), and two NF-kB enhancer elements at positions −104 to −81 ( 14 ). The core positive strand promoter is composed of three Spl binding sites located at positions −78 through −47 and a TATA box at positions −28 to −24 ( 15 , 16 ). A 59-base pair region which confers transcriptional transactivating potential to the core promoter elements by the viral Tat protein is located within the R region of the LTR at positions +1 to +59 ( 17 , 18 ). This transactivating region (TAR) folds into alternate RNA stem-loop structures that are found in the 5′ termini of all viral transcripts. The TAR element mediates a substantial increase in transcriptional initiation and elongation through interactions with the viral Tat protein and other cellular factors ( 3 , 19 – 22 ).

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